Sinteza, karakterizacija i biološka aktivnost kompleksa Co(III) sa kondenzacionim proizvodom 2-acetilpiridina i dihidrazida malonske kiseline

Abstract

U ovom radu opisana je sinteza, karakterizacija i biološka aktivnost kompleksa kobalta(III) sa kondenzacionim proizvodom 2-acetilpiridina i dihidrazida malonske kiseline. TakoĊe, opisana je sinteza i karakterizacija organske soli 2,2-{1,1-[2,2-oksalilbis(hidrazin-2-il-1-iliden)]dietilidin}dipiridinijum diperhlorata dihidrata. Sintetisane supstance su okarakterisane u rastvoru NMR spektroskopijom, a u čvrstom stanju rendgenskom strukturnom analizom. U dobijenom kompleksu kobalta(III) kompleksni katjon [Co2L22]2+ poseduje dinuklearnu strukturu dvostrukog heliksa u kojoj je svaki Co(III) centar koordinovan preko dve tridentatne N2O helatne jedinice iz oba H2L2 liganda, formirajući oko oba Co(III) oktaedarsko okruţenje. Citotoksični potencijal kompleksa je uočen na ćelijama epitelnog kancera dojke (MDA-361) sa IC50 vrednošću 50,9 μM. Slaba citotoksična aktivnost je uočena na EA.hy926 i B16 ćelijskim linijama sa IC50 vrednošću 258 μM odnosno 221 μM. Kompleks pokazuje slabu citotoksičnost na MRC-5 ćelije. H2L2 ligand pokazuje umerenu aktivnost na sve ćelijske linije osim na K562 i MDA-361 ćelije, a najbolja aktivnost je uočena na ćelijama mišijeg melanoma B16. K562 ćelije su rezistentne na kompleks i ligand, štaviše kompleks stimuliše njihov rast. Ćelije epitelnog kancera dojke MDA-361 su korišćene u eksperimentima u kojima je ispitan uticaj kompleksa [Co2L22](NO3)2•2H2O•0,5C2H5OH na progresiju ćelijskog ciklusa i indukciju ćelijske smrti. Posle 24 h inkubacije ćelija tretiranih kompleksom koncentracije IC50 i 2 x IC50 nije došlo do značajne promene u zastupljenosti ćelija u G1, S i G2/M fazi. Uočen je jasan sub-G1 pik koji potiče od hipodiploidnih ćelija koje sadrţe fragmentisane molekule DNA što ukazuje na ćelije u kasnoj fazi apoptoze. Apoptotički potencijal kompleksa [Co2L22]2+ na MDA-361 ćelijama je ispitan i pomoću Annexin V-FITC / PI testa. Ispitivani kompleks dovodi do značajanog povećanja procenta ćelija u ranoj fazi apoptoze (28,18 %) u odnosu na kontrolne ćelije (5,64 %). Interakcije kompleksa sa DNA iz telećeg timusa (CT-DNA) dovode do plavog pomeranja i hiperhromizma u UV-Vis spektrima. Vrednost konstante vezivanja Kb=4.2×105 M−1 ukazuje na vezivanje kompleksa u brazdi DNA.

In this work the synthesis, characterization and biological activity of cobalt(III) with the condensation product of 2-acetylpyridine and malonic acid dihydrazide are presented. Furthermore the synthesis and characterization of organic salts 2,2-{1,1-[2,2-oxalylbis(hydrazin-2-yl-1-ylidene)]diethylidyne}dipyridinium bis(perchlorate) dehydrate have been shown. Obtained substances were characterized in solution by NMR spectroscopy and in solid-state by the X-ray structure analysis. The complex cation [Co2L22]2+ possesses a dinuclear double-stranded helicate structure in which each of Co(III) center is coordinated by two tridentate N2O chelating units from different ligands. Each Co(III) center thus are in a six-coordinated pseudooctahedral environment. Cytotoxic potential was observed for the complex on the epithelial breast cancer (MDA-361) cell line, where the IC50 was 50.9 μM. The weak cytotoxic activity of the complex was observed with EA.hy926 and B16 cell line, reaching IC50 values at 258 μM and 221 μM, respectively. The complex revealed low cytotoxicity against MRC-5 cells. The corresponding H2L2 ligand also showed moderate cytotoxic activity against all cell lines, except K562 and MDA-361 cells, with the highest potency on the murine melanoma B16 cell line. K562 cells were resistant to both complex and ligand, the complex even stimulating cells to grow. Epithelial breast cancer (MDA-361) cell lines were used in the experiments where we examinated influence of the complex [Co2L22](NO3)2·2H2O·0,5C2H5OH on cell cycle progression and induction of cell death. After 24 h of incubation of cells treated with the concentration of IC50 and 2× IC50, the investigated complex gave no significant changes in the percentage of cells in the G1, S and G2/M phases. There is a clearly determined sub-G1 peak in the histograms of treated hypodiploid cells, containing fragmentized DNA molecule, indicating that such cells are in late apoptotic phase. Apoptosis of MDA-361 cells treated with investigated complex was evaluated by Annexin V–FITC / PI test. The investigated complex induced apoptosis, the early apoptotic cells comprising 28.18%, compared to 5.64% of control cells in the same phase. The interaction of the complex with calf thymus DNA (CT-DNA) was monitored by blue shift and hyperchromism in the UV–vis spectra. The observed intrinsic binding constant (Kb=4.2×105 M−1) together with structural analysis of the complex indicate the groove binding.